Life and development of all organisms are determined by molecular interactions, e.g. between DNA and proteins, proteins and proteins, or proteins and small molecules. Among these, protein-protein interactions play an especially important role, for example with the interactions between antibodies and antigens, receptors and peptide- or protein-hormones, enzymes and substrates or inhibitors. Many of the best-selling drugs either act by targeting proteins or are proteins. In addition, many molecular markers of disease, which are the basis of diagnostics, are proteins.
The development of techniques and reagents for high throughput protein analysis has been of great interest. In particular, the increasing knowledge of DNA sequence in organisms of interest has spurred interest in protein expression analysis. There is now a rapidly growing awareness of just how important proteomics is to understand and organize the human genome. Information about the complement of proteins present in a cell is a key to accelerate the discovery of medically important proteins and the genes from which they derive.
Genomics establishes the relationship between gene activity and particular diseases. However most disease processes are manifested not at the level of genes, but at the protein level. There is often a poor correlation between the level of activity of different genes and the relative abundance of the corresponding proteins. Also a protein and its post-translational modifications are not directly encoded for by the same gene, therefore the complete structure of individual proteins cannot be determined by reference to the gene alone.
Assays directed towards protein binding can be used for the quantitation of protein expression; the determination of specific interactions; to determine the presence of ligands for a protein, and the like. Methods of quantitating proteins in a sample by determining binding to a cognate antibody are known in the art.
For example, solid-phase radioimmunoassay (RIA) of antigens or antibodies in a serum sample are well known. Catt et al. have reported such techniques on the surface of plastic tubes (U.S. Pat. No. 3,646,346) and plastic discs (J. Lab. & Clin. Med., 70: 820 (1967). In such techniques, an excess of specific antibody is first adsorbed to a support surface. Then, the sample to be assayed is immunologically reacted with such surface in a sandwich or competitive binding technique. In the competitive binding technique, illustrated in U.S. Pat. No. 3,555,143, the concentration of antigen to be determined and a known quantity of radioactively tagged antigen are immunologically reacted with the antibody-adsorbed surface. The labeled antigen bound to the antibody on the surface is then quantitated to determine indirectly the total quantity of antigen in the original sample. In the sandwich technique, serum containing an unknown concentration of antigen is immunologically reacted with the antibody-containing surface. Then in a following step, the bound antigen is incubated with labeled antibody and the amount of immunologically bound, labeled antibody is subsequently measured.
The development of high-throughput, parallel systems for protein analysis are of great interest, particularly where the analysis can use small amounts of material for analysis. Preferably such systems provide for the use of complex molecules with high binding affinity for their ligands, such as antibodies, protein receptors, and the like.